LOCALIZATION OF THE DISULFIDE BOND INVOLVED IN POSTTRANSLATIONAL PROCESSING OF GLYCOSYLASPARAGINASE AND DISRUPTED BY A MUTATION IN THE FINNISH-TYPE ASPARTYLGLYCOSAMINURIA

The heavy chain of human glycosylasparaginase (N-4-(beta-N-acetylglucosaminyl)-L-asparaginase (EC 3.5.1.26)) has five cysteinyl residues (Cys-61, Cys-64, Cys-69, Cys-163, and Cys-179). A Cys-163 to serine substitution due to a point mutation in the glycosylasparaginase gene causes the most common disorder of glycoprotein degradation, the Finnish-type aspartylglycosaminuria. To localize the potential disulfide bonds, the isolated heavy chain of human leukocyte glycosylasparaginase was treated with the enzyme alpha-chymotrypsin, and the resulting peptides were separated by high performance liquid chromatography prior to and after reduction and S-carboxymethylation. The peptide containing the Cys-163 residue and the peptide to which it is connected with a disulfide were structurally characterized by mass spectrometry. The disulfide bond crucial for catalytic activity, subunit processing, and biological transport of glycosylasparaginase was located close to the carboxyl terminus of the heavy chain at positions 163 and 179.